The capability of spermatozoa to directly influence maternal gene expression is already established. Indeed, some of the changes induced by spermatozoa may have a direct functional importance in the pre-conceptional period. Although the mechanisms underlying these sperm-maternal interactions are not well characterized, it is possible that they could involve ligands that are released from the spermatozoa. This study therefore aimed to test whether physical contact between bovine spermatozoa and bovine oviductal epithelial cells (BOECs) is a prerequisite for spermatozoa-induced gene expression changes. We used two co-culture models: a contact co-culture model in which spermatozoa interact directly with BOECs, and a non-contact co-culture model in which an insert with the pore size of 0.4 μm was placed between spermatozoa and BOECs. Messenger RNA sequencing analysis of BOECs by RNA-seq revealed ten differentially expressed genes in contact system and 108 differentially expressed genes in the non-contact system after 10 h of co-culture. Retinol metabolism pathway and ovarian steroidogenesis pathway were significantly enriched in the non-contact co-culture system. Q-PCR analysis revealed that transcriptional responses can be rapid, with increased expression of four genes (DHRS3, CYP1B1, PTGS2, and ATF3) detectable within just 90 min of co-incubation, but with expression levels highly dependent on the type of co-culture system. The findings from our study demonstrate that direct contact with spermatozoa is not necessary to induce changes in gene expression of oviductal epithelial cells, suggesting that spermatozoa may be able to signal to maternal tissues in advance of their arrival.

已经建立了精子直接影响母体基因表达的能力。确实，精子诱导的某些变化在受孕前期可能具有直接的功能重要性。尽管这些精子与母体之间相互作用的机制尚未得到很好的表征，但它们可能涉及从精子中释放出来的配体。因此，本研究旨在检验牛精子和牛输卵管上皮细胞（BOEC）之间的物理接触是否是精子诱导的基因表达变化的前提。我们使用了两种共培养模型：一种接触共培养模型，其中精子直接与BOEC相互作用；以及一种非接触共培养模型，其中将孔径为0.4μm的插入物置于精子和BOEC之间。通过RNA-seq对BOEC进行信使RNA测序分析，共培养10 h后，接触系统中有10个差异表达基因，非接触系统中有108个差异表达基因。在非接触式共培养系统中，视黄醇代谢途径和卵巢类固醇生成途径明显丰富。Q-PCR分析显示转录反应可以快速进行，其中四个基因的表达增加（DHRS3，CYP1B1，PTGS2和ATF3）在共孵育的90分钟内即可检测到，但表达水平高度依赖于共培养系统的类型。我们研究的结果表明，与精子直接接触并不是诱导输卵管上皮细胞基因表达变化的必要条件，这表明精子可能能够在到达母体组织之前向母体组织发出信号。